Targeted at learning about inflammatory eye diseases, this project focused during FY 2006 on three topics: (1) the mechanisms whereby pertussis toxin stimulates naive T-lymphocytes to acquire effector cell features and initiate severe immunopathogenic processes ; (2) the expression of anti-inflammatory molecules in the eye; (3) suppression of immune-mediated ocular inflammation by synthetic copolymers.[unreadable] [unreadable] Topic 1.[unreadable] In a study carried out in FY 2005 we made the observation that microbial products, that are ligands for toll-like receptors (TLRs) on antigen presenting cells, are capable of triggering pathogenic autoimmune processes in which non-pathogenic naive T-lymphocytes specific toward eye-specific antigens acquire features of ?effector? cells and initiate ocular inflammation. Of the seven TLR ligands tested in this study, pertussis toxin was found to be the most active agent by far, triggering extremely severe inflammation in eyes of treated mice. Further analysis of this observation in FY 2006 revealed that the superior activity of pertussis toxin is related to its capacity to affect the naive cells by three mechanisms: (i) stimulation of vigorous proliferation by these cells; (ii) polarization of the T-cells toward the highly immunopathogenic ?type 1? response, characterized by production of interferon gamma and (iii) modification of these cells? surface markers, to make them capable of efficiently invading the target organ, the eye. By all three parameters, treatment with pertussis toxin exceeded by far the activity of the other six tested TLR ligands, suggesting that these effects of pertussis toxin on the naive recipient cells are major mechanisms for the severe inflammatory inflammation seen in mice treated with this molecule. Futhermore, our data shed new light on the mechanism whereby pertussis toxin functions as an adjuvant, enhancing a variety of pathogenic autoimmune processes.[unreadable] [unreadable] Topic 2.[unreadable] Ocular tissues are highly susceptible to the detrimental effects of inflammation and, therefore, the eye is protected from these effects by a mechanism (?ACAID?) that directs the immune system toward immunotolerance rather than immune response, as well as by expressing several immunosuppressive molecules, such as TGF beta. Recent publications have identified two new molecules that exhibit anti-inflammation activities, ?GC binding protein (GC-BP)? (Kim et al., Immunity, 2004, 21:643) and ?Triggering receptor expressed on myeloid cells-2 (TREM-2)?(Takahashi et al., J. Exp. Med., 2005, 201:647). In our study, we detected both molecules in normal mouse eyes, as well as in cell lines of human retinal pigment epithelium and Muller cells. Mouse eyes developing experimental autoimmune uveoretinitis (EAU) exhibited a remarkable increase in expression of TREM-2, but showed no significant change in the level of GC-BP, in line with the observation that this molecule undergoes functional upregulation during inflammation, rather than increase in concentration. Our results suggest that both TREM-2 and GC-BP contribute to the protective anti-inflammatory mechanism of the eye. [unreadable] [unreadable] Topic 3.[unreadable] ?Copaxone?, also known as ?Glatiramer acetate?, is a synthetic copolymer that was found to inhibit an animal model for multiple sclerosis and is currently in use for treatment of this disease. We have previously reported that Copaxone also partially inhibits the development of EAU in mice of the B10.A strain when administered together with the retinal antigen in the adjuvant emulsion used for disease induction (Zhang et al., J. Neuroimmunol., 2000, 103:189). The study was expanded in FY 2006, yielding the following main findings: (i) A new copolymer, designated ?YFAK?, that is composed of amino acids different from those of Copaxone, was found to be equal or superior to Copaxone in its capacity to inhibit EAU induction; (ii) the suppressive effects of the two copolymers were particularly apparent when tested in the EAU model with another mouse strain, B10.RIII; (iii) While being effective in inhibiting EAU when administered in emulsion together with the uveitis-inducing antigen, the two copolymers had little or no effect when given separately from the antigen-adjuvant emulsion. This observation suggests that the copolymers? inhibitory effect is mainly by these molecules' capacity to compete with the uveitogenic antigen for the combining sites on the major histocompatibility complex (MHC) molecules on antigen presenting cells.